New genes of Xanthomonas citri subsp citri involved in pathogenesis and adaptation revealed by a transposon-based mutant library

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Pathogenesis II: Fungal responses to host responses: interaction of host cells with fungi

Most of our knowledge concerning the virulence determinants of pathogenic fungi comes from the infected host, mainly from animal models and more recently from in vitro studies with cell cultures. The fungi usually present intra- and/or extracellular host-parasite interfaces, with the parasitism phenomenon dependent on complementary surface molecules. Among living organisms, this has been characterized as a cohabitation event, where the fungus is able to recognize specific host tissues acting as an attractant, creating stable conditions for its survival. Several fungi pathogenic for humans and animals have evolved special strategies to deliver elements to their cellular targets that may be relevant to their pathogenicity. Most of these pathogens express surface factors that mediate binding to host cells either directly or indirectly, in the latter case binding to host adhesion components such as extracellular matrix (ECM) proteins, which act as 'interlinking' molecules. The entry of the pathogen into the host cell is initiated by fungal adherence to the cell surface, which generates an uptake signal that may induce its cytoplasmic internalization. Once this is accomplished, some fungi are able to alter the host cytoskeletal architecture, as manifested by a rearrangement of microtubule and microfilament proteins, and this can also induce epithelial host cells to become apoptotic. It is possible that fungal pathogens induce modulation of different host cell pathways in order to evade host defences and to foster their own proliferation. For a number of pathogens, the ability to bind ECM glycoproteins, the capability of internalization and the induction of apoptosis are considered important factors in virulence. Furthermore, specific recognition between fungal parasites and their host cell targets may be mediated by the interaction of carbohydrate-binding proteins, e.g., lectins on the surface of one type of cell, probably a parasite, that combine with complementary sugars on the surface of host-cell. These interactions supply precise models to study putative adhesins and receptor-containing molecules in the context of the fungus-host interface. The recognition of the host molecules by fungi such as Aspergillus fumigatus, Paracoccidioides brasiliensis and Histoplasma capsulatum, and their molecular mechanisms of adhesion and invasion, are reviewed in this paper.

Traumatic subdural hygromas: Proposed pathogenesis based classification

Background: Traumatic subdural hygroma (TSHy) is an accumulation of cerebrospinal fluid (CSF) in the subdural space after head injury. It appears to be relatively common, but its onset time and natural history are not well defined. Considered a benign epiphenomenon of trauma, the pathogenesis of TSHy is still unclear and many questions remain unanswered. This study adds to the information on TSHy, and proposes a classification based on pathogenesis.Methods: Thirty-four consecutive adult patients with TSHy were analyzed for clinical evolution and serial CT scan, during a period of several months. TSHy diagnosis was based on published CT scan criteria of hypodense subdural collection after trauma, without enhancement and neomembrane, with a minimum distance of 3 mm between the skull and brain. Ventricle size was analyzed by calculating the bicaudate index (BCI). For comparison, the BCI was measured from CT scan at three moments: admission, at time of TSHy diagnosis, and from last CT scan.Results: There were 34 patients, aged between 16 and 85 years (mean 40), half of them were below 40 years. Road traffic crashes were the main cause of head injury. The mean time for hygroma diagnosis was 9 days. Twenty-one patients (61.8%) underwent conservative treatment for TSHy and 13 (38.2%), surgical treatment. TSHy are early lesions and can be detected in the first 24 hours after trauma, usually as small subdural effusion (SSEff). Based on clinical and CT scan findings, we divided the 34 patients into 3 groups, (Ia and Ib) without evident mass effect and (II) with evident mass effect. Group Ia includes patients without ventricle dilation; Ib, patients with associated ventricle dilations.Conclusions: SSEff detected in the first 24 hours posttrauma in our series evolved into TSHy suggesting that this is an early lesion; all THSy were divided in three groups according to the pathophysiologic mechanism. These three groups probably represent a continuum of CSF absorption impairment. Group la represents what most authors consider a simple hygroma, with no impairment on CSF absorption. Group Ib represent the external hydrocephalus form with various degrees of CSF imbalance, and group II were the cases presenting marked mass effect.

HIV, EBV and KSHV: Viral cooperation in the pathogenesis of human malignancies

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The heminested RT-PCR for the study of rabies virus pathogenesis

The aim of the present trial was to evaluate the heminested RT-PCR for the study of rabies virus distribution in mice inoculated experimentally. Inoculation was by the intramuscular route in 150 mice, using the dog street rabies virus. Groups of five animals were killed at different times. Fragments of different organs were collected and the material was tested by Fluorescent Antibody Test (FAT) and heminested RT-PCR (hn RT-PCR). Positive results were obtained beginning on the 10th day after inoculation in the brain, spinal cord, salivary gland, limbs, lungs, liver, spleen, urinary bladder, tongue and right kidney. Hn RT-PCR was shown to be more efficient for the study of rabies virus distribution in different tissues and organs. (C) 2004 Elsevier B.V.. All rights reserved.

Influence of Antiplatelet Drugs in the Pathogenesis of Experimental Periodontitis and Periodontal Repair in Rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

In Vivo Models for Measuring Placental Glutatione-S-transferase (GST-P 7-7) Levels: A Suitable Biomarker for Understanding Cancer Pathogenesis

The Glutatione-S-transferases (GSTs) comprise a family of enzymes closely associated with the cell detoxification of xenobiotics. GSTs exist as homo- or heterodimers and have been grouped into at least seven distinct classes. The main function of GSTs is to catalyze the conjugation of reduced glutathione (GSH) to an electrophilic site of a broad range of potentially toxic and carcinogenic compounds, thereby making such compounds less dangerous and enabling their ready-excretion. Placental GST, known as GST-P 7-7, is the main isoform found in normal placental tissue and comprises 67% of the total GST concentration in this tissue. During development, GST-P 7-7 decreases in concentration and is absent in adult tissues. Interestingly, GST-P 7-7 expression has been detected in adult tissues after exposure to carcinogenic agents in several experimental test systems, being considered a reliable biomarker of exposure and susceptibility in early phases of carcinogenesis. In this article, we review a series of studies involving GST-P 7-7 expression as a suitable tool for understanding cancer pathogenesis, especially cancer risk.

Inheritance of resistance to powdery mildew in pea and pathogenesis-related aspects

A herança da resistência ao oídio na cultivar de ervilha MK-10 e alguns aspectos histológicos da infecção foram estudados. Para o estudo da herança, as gerações F1, F2, retrocuzamentos e geração F3 de MK-10 com duas populações suscetíveis foram avaliadas. Nas avaliações histológicas observou-se a porcentagem de conídios germinados, porcentagem de conídios que formaram apressório, porcentagem de conídios que estabeleceram colônia e número de haustórios por colônia. Para comparar as razões de segregação obtidas no estudo da herança da resistência, adotou-se o teste do Qui-quadrado (X²) e para os dados das análises histológicas, utilizou-se o teste Tukey a 5% de probabilidade. Concluiu-se que a resistência de MK-10 ao oídio é devida a um par de alelos recessivos e que a resistência é expressa na fase de pré-penetração, completada por uma morte celular localizada pós-penetração, característica da presença do par de alelos recessivos er1er1.

Implications of genetic variability of Trypanosoma cruzi for the pathogenesis of Chagas disease

O Trypanosoma cruzi, agente etiológico da doença de Chagas, apresenta elevado grau de variabilidade genética intra-específica, com possíveis implicações na forma clínica da doença, como o desenvolvimento de cardiopatia, do megaesôfago e do megacólon de forma isolada ou em associação. Este tropismo tecidual envolvido na patogênese da doença não está totalmente esclarecido. Assim, nesta revisão são abordados alguns aspectos referentes à diversidade genética dos parasitas isolados, às formas clínicas da doença de Chagas, ao processo de infecção do parasita na célula hospedeira e resposta imune. Outros aspectos também são enfocados, como os fatores imunossupressivos liberados pelo parasita que atuam na regulação das respostas imunes, a inibição da apoptose da célula hospedeira, assim como da patogênese do megaesôfago chagásico que pode estar relacionada à interação hospedeiro- parasita e sua associação com risco aumentado para o desenvolvimento do carcinoma epidermóide do esôfago. Porém, apesar dos avanços no entendimento desta doença, ainda não é possível estabelecer o verdadeiro perfil da variabilidade genética do parasita com a forma clínica da doença de Chagas.

HAEMONCHUS PLACEI IN CALVES - EFFECTS OF DIETARY-PROTEIN AND MULTIPLE EXPERIMENTAL-INFECTION ON WORM ESTABLISHMENT AND PATHOGENESIS

An experiment was conducted to examine the influence of dietary protein and immunisation on parasite establishment and pathogenesis of Haemonchus placei in calves. Four groups of 4-6-month-old worm-free calves (n=4) were given a low protein diet (LP) containing 213 g crude protein (CP) per head per day or a high-protein diet (HP) containing 469 g per head per day CP. Five weeks later, calves in one of the two groups of each dietary treatment were given 50 000 H. placei infective larvae (L(3)). Twenty-five days later, infection in these groups was terminated by dosing with oxfendazole, This immunisation process was repeated 4 days later. Four days after termination of the second immunisation all calves were challenged with 100 000 L(3). Five weeks later, all calves were slaughtered for abomasal worm counts. Worm establishment was lower in the immunised groups; however, only the HP-I group showed a significant reduction (P < 0.05). All calves gained weight during the first 13 week period, and after challenge the non-immunised groups lost weight, independent of the level of protein in the diet (P < 0.05), Packed cell volume values for all treatments only dropped after challenge (P < 0.05) and the HP-immunised group presented values significantly higher when compared with the other treatments, All calves were hypoproteinaemic and hypoalbuminaemic at the end of the experiment, regardless of the treatment. Immunised calves showed a normocytic normochromic anaemia, while the non-immunised groups presented a microcytic normochromic anaemia.

BINDING OF PARACOCCIDIOIDES-BRASILIENSIS TO LAMININ THROUGH SURFACE GLYCOPROTEIN GP43 LEADS TO ENHANCEMENT OF FUNGAL PATHOGENESIS

Extracellular matrix protein laminin binds specifically to yeast forms of Paracoccidioides brasiliensis and enhances adhesion of the fungus to the surface of epithelial Madin-Darby canine kidney cells in vitro. Immunoblotting of fungal extracts showed that the gp43 glycoprotein is responsible for adhesion. This was confirmed by binding assays using purified gp43, with a K-d of 3.7 nM. The coating of P. brasiliensis yeast forms with laminin before injection into hamster testicles enhanced the fungus virulence, resulting in a faster and more severe granulomatous disease. These results indicate that interaction of fungi with extracellular matrix elements may constitute a basis for the evolution of fungal infection toward regional spreading and dissemination.

Experimental alcoholism and pathogenesis of prostatic diseases in UChB rats

Previous studies have shown that long-term alcohol treatment has negative effects on prostatic stromal-epithelial interaction. Thus, the aim of the present study was to analyze the histochemical, immunohistochemical and ultrastructural alterations that occur in the prostatic stroma and epithelium of rats submitted to chronic alcohol ingestion and alcohol abstinence, as well as to establish the relationship between these changes and prostatic diseases. Thirty male rats (10 Wistar and 20 UChB rats) were divided into three experimental groups: the control group received tap water, the alcoholic group received ethanol diluted to 10 degrees G.L. for 150 days, and the abstinent group received the same liquid diet as the alcoholic group up to 120 days of treatment and only tap water for 30 days thereafter. At the end of treatment, all animals were sacrificed and the ventral lobe of the prostate was removed and processed for histochemical, immunohistochemical and ultrastructural analyses. In addition, plasma testosterone levels were measured. The results showed, prostatic intraepithelial neoplasia, infolding of the epithelium towards the stroma, stromal hypertrophy and the presence of inflammatory cells in alcoholic animals. In the abstinent group, alterations were noted mainly in the stromal area. In conclusion, ethanol triggers alterations in prostatic epithelial and stromal compartments, affecting the stromal microenvironment and predisposing the organ to pathological processes. (C) 2006 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.

Monoclonal antibodies against the 43,000 da glycoprotein from Paracoccidioides brasiliensis modulate laminin-mediated fungal adhesion to epithelial cells and pathogenesis

The surface glycoprotein gp43, a highly immunogenic component of Paracoccidioides brasiliensis, is used in the serodiagnosis of paracoccidioidomycosis (PCM) and has recently been shown to specifically bind the extracellular matrix protein laminin, Binding to laminin induces the increased adhesion of the fungus to epithelial cells; a hamster testicle infection model has shown that the gp43-dependent binding of fungal cells to laminin enhances their pathogenicity in vivo. We report on the production and characterization of 12 monoclonal antibodies against the gp43 that recognize peptide sequences in the molecule detecting at least three different epitopes as well as different isoforms of this antigen. MAbs interfered in the fungal pathogenicity in vivo either by inhibiting or enhancing granuloma formation and tissue destruction, Results suggest that P. brasiliensis propagules may start infection in man by strongly adhering to human lung cells, Thus, laminin-mediated fungal adhesion to human lung carcinoma (A549) cells was much more intense than to Madin-Darby canine kidney cells (MDCK), indicating differences in binding affinity, Subsequent growth of fungi bound to the lung cells could induce the granulomatous inflammatory reaction characteristic of PCM. Both steps are greatly stimulated by laminin binding in infective cells expressing gp43.